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The in vivo endothelial cell translatome is highly heterogeneous across vascular beds

ABSTRACT: This SuperSeries is composed of the SubSeries listed below.

ORGANISM(S): Mus musculus Homo sapiens

PROVIDER: GSE138630 | GEO | 2019/10/10

REPOSITORIES: GEO

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The in vivo endothelial cell translatome is highly heterogeneous across vascular beds.

Project description:Endothelial cells (ECs) are highly specialized across vascular beds. However, given their interspersed anatomic distribution, comprehensive characterization of the molecular basis for this heterogeneity in vivo has been limited. By applying endothelial-specific translating ribosome affinity purification (EC-TRAP) combined with high-throughput RNA sequencing analysis, we identified pan EC-enriched genes and tissue-specific EC transcripts, which include both established markers and genes previously unappreciated for their presence in ECs. In addition, EC-TRAP limits changes in gene expression after EC isolation and in vitro expansion, as well as rapid vascular bed-specific shifts in EC gene expression profiles as a result of the enzymatic tissue dissociation required to generate single-cell suspensions for fluorescence-activated cell sorting or single-cell RNA sequencing analysis. Comparison of our EC-TRAP with published single-cell RNA sequencing data further demonstrates considerably greater sensitivity of EC-TRAP for the detection of low abundant transcripts. Application of EC-TRAP to examine the in vivo host response to lipopolysaccharide (LPS) revealed the induction of gene expression programs associated with a native defense response, with marked differences across vascular beds. Furthermore, comparative analysis of whole-tissue and TRAP-selected mRNAs identified LPS-induced differences that would not have been detected by whole-tissue analysis alone. Together, these data provide a resource for the analysis of EC-specific gene expression programs across heterogeneous vascular beds under both physiologic and pathologic conditions.

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The in vivo endothelial cell translatome is highly heterogeneous across vascular beds (HUVEC RNAseq)

Project description:The interspersed anatomic distribution of endothelial cells (ECs) and their dependence on microenvironmental cues have been major challenges in studying EC function and response to (patho)physiological stimuli in vivo. Combining EC-specific translating ribosome affinity purification (EC-TRAP) with RNA sequencing provides an accurate in vivo snapshot of tissue-specific EC expression profiles, and maintains a high degree of sensitivity to detect low abundant transcripts while limiting changes in gene expression profiles due to enzymatic tissue dissociation required to generate single cell suspensions for FACS sorting or single cell analysis. In vitro HUVEC data was obtained to ensure short-term cycloheximide treatment, as used for in vivo experiments, did not alter global gene expression.

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Project description:The interspersed anatomic distribution of endothelial cells (ECs) and their dependence on microenvironmental cues have been major challenges in studying EC function and response to (patho)physiological stimuli in vivo. Combining EC-specific translating ribosome affinity purification (EC-TRAP) with RNA sequencing provides an accurate in vivo snapshot of tissue-specific EC expression profiles, and maintains a high degree of sensitivity to detect low abundant transcripts while limiting changes in gene expression profiles due to enzymatic tissue dissociation required to generate single cell suspensions for FACS sorting or single cell analysis. In addition to demonstrating EC heterogeneity under physiologic conditions, the in vivo host response to lipopolysaccharide (LPS) revealed the induction of expression programs associated with a native defense response, with marked differences across vascular beds.

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